1. Field of the Invention
The present invention relates to a method of producing ethanol using continuous culture in which recombinant microorganisms having xylose-metabolizing ability is used; and a continuous culture apparatus.
2. Description of Related Art
Main saccharides contained in lignocellulose include glucose constituting cellulose and xylose constituting hemicellulose. When lignocellulose is chemically or enzymatically decomposed, a saccharified composition containing monosaccharides such as glucose and xylose as major components is obtained. In order to industrially produce a useful material from lignocellulose, microorganisms which can be fermented with high yield and high productivity by efficiently using saccharides contained in the saccharified composition; and a production process are required.
For example, genes encoding xylose isomerase (XI), xylose reductase (XR), and xylitol dehydrogenase (XDH) are transferred to a yeast having high ethanol fermentation ability such as Saccharomyces cerevisiae to impart xylose utilizing ability to a yeast. With this yeast, ethanol can be produced by efficiently using saccharides derived from lignocellulose.
In general, as a method of producing by fermentation a material using microorganisms, batch culture, continuous culture, and feeding culture (fedbatch culture) are known. In batch culture, a new culture medium is prepared for each process, microorganisms are inoculated into the prepared culture medium, and another culture medium is not added until the end of the culture. In batch culture, the quality of each culture varies, but the risk of contamination can be dispersed and reduced. In continuous culture, a culture medium is supplied to a culture system at a constant rate, and concurrently, a culture fluid is extracted from the culture system at the same rate. Continuous culture is characterized in that a constant culture environment is easily maintained and productivity is stable. In feeding culture, a culture medium or a specific component of a culture medium is added to a culture system during the culture, and the obtained product is not extracted from the culture system until the end of the culture.
In particular, in regard to continuous culture, Japanese Patent Application Publication No. 2009-065970 (JP 2009-065970 A) discloses a method of producing a chemical product by continuous fermentation in which high productivity can be stably maintained over a long period of time under simple operating conditions. In the method disclosed in JP 2009-065970 A, a fermentation culture fluid is filtered through a porous film, and microorganisms and the like are extracted from the fermentation culture fluid to the outside of the system. However, in this method, when a lignocellulose-based material is used, the clogging of the porous film occurs, and there is a problem in that high cost is required to solve the clogging.
Japanese Patent Application Publication No. 2010-227074 (JP 2010-227074 A) discloses a method of continuously producing ethanol by alcohol fermentation in which a yeast is used with a starchy material as a substrate. In the method disclosed in JP 2010-227074 A, continuous fermentation conditions including fermentation temperature, substrate supply rate, fermentation liquid extraction rate, air flow rate, and yeast concentration are controlled to specific values, and the ethanol concentration in the fermentation liquid is maintained to be a predetermined value or lower. According to the method disclosed in JP 2010-227074 A, the production of ethanol can be efficiently continued without additional supply of a yeast during continuous fermentation. However, in the continuous culture method disclosed in JP 2010-227074 A, when saccharides derived from lignocellulose are used as a substrate, there is a problem in that ethanol cannot be efficiently produced.